Uninary tract infection (UTI) is one of the most common bacterial infections in developed countries; hospital acquired UTI is the most frequent nosocomial infection. Escherichia coli is the most frequent cause of both community- and hospital-acquired UTI. Uropathogenic E. coli possess virulence associated factors not as frequently found among fecal E. coli isolates. The long term goals of this of this research are to identify new methodologies for the prevention and treatment of UTI by E. coli, by acquiring a more thorough understanding of its pathogenic mechanisms. The immediate objectives of this project are to identify those bacterial factors which are both necessary and sufficient for E. coli to colonize the mouse urinary tract in an ascending model of pyelonephritis. The results of our experiments have demonstrated the necessities for P- and type-1 fimbriae, and the polysaccharide antigens o and K in the capacity of E. coli to colonize the mouse UT. In this application we propose to clone the genes for O and k antigen synthesis to further quantify their roles in virulence. Recombinant plasmids encoding the enzymes required for the synthesis of these polysaccharides will be transferred to a nonpathogenic laboratory E. coli strain, and the various isogenic transformants will be compared with respect to (1) resistance to the bactericidal effects of serum, and to ingestion by phagocytes, and (2) capacity to colonize the UT of mice. Mouse infection studies will be performed by Dr. Catharina Svanborg-Eden, University of Goteborg, Sweden. Furthermore, we have discovered that P-fimbriae are not identical with respect to receptor recognition. Recombinant plasmids encoding the three different types of P-fimbriae will be constructed (only one remains to be subcloned) and transferred to a nonpathogenic fecal isolate of E. coli to test the relative capacities of each type of infect mice.